LOCUS       sequence-227761-       10352 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses dCas9-2xNLS-EGFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10352)
  AUTHORS   Nelles DA, Fang MY, O'Connell MR, Xu JL, Markmiller SJ, Doudna JA, 
            Yeo GW
  TITLE     Programmable RNA Tracking in Live Cells with CRISPR/Cas9.
  JOURNAL   Cell. 2016 Mar 16. pii: S0092-8674(16)30204-5. doi: 
            10.1016/j.cell.2016.02.054.
  PUBMED    26997482
REFERENCE   2  (bases 1 to 10352)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell. 2016 
            Mar 16. pii: S0092-8674(16)30204-5. doi: 
            10.1016/j.cell.2016.02.054."
COMMENT     Sequence Label: pCDNA3.1- dCas9-2xNLS-EGFP
FEATURES             Location/Qualifiers
     source          1..10352
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(727..753,754..756,757..3243,3244..3246,3247..4830)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             727..4830
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             4834..4860
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4840..4866
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             4879..4899
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4906..4926
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(4930..4932,4933..5646)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(4930..4932,4933..5643)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(5233..5252)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5580..5601
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(5709..5726)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5715..5939
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      5985..6413
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6072..6091)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6282..6303
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6422..6442)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6427..6756
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6607..6742
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     6669..6688
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             6823..7617
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6877..6896)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7487..7506
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    7791..7912
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7828..7847)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7882..7901
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7961..7977)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7961..7977)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7974..7996)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7985..8001
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(8009..8015,8016..8033,8034..8039))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8054..8075
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8192..8209)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8363..8951)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8443..8462)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9122..9913,9914..9982))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     9745..9764
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9983..10087)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(10162..10181)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctg
      661 gctagcgttt aaacgggccc tctagactcg agcggccgcc actgtgctgg atatctgcag
      721 agaaccatgg acaagaagta cagcatcggc ctggccatcg gcaccaactc tgtgggctgg
      781 gccgtgatca ccgacgagta caaggtgccc agcaagaaat tcaaggtgct gggcaacacc
      841 gaccggcaca gcatcaagaa gaacctgatc ggcgccctgc tgttcgacag cggagaaaca
      901 gccgaggcca cccggctgaa gagaaccgcc agaagaagat acaccagacg gaagaaccgg
      961 atctgctatc tgcaagagat cttcagcaac gagatggcca aggtggacga cagcttcttc
     1021 cacagactgg aagagtcctt cctggtggaa gaggataaga agcacgagcg gcaccccatc
     1081 ttcggcaaca tcgtggacga ggtggcctac cacgagaagt accccaccat ctaccacctg
     1141 agaaagaaac tggtggacag caccgacaag gccgacctgc ggctgatcta tctggccctg
     1201 gcccacatga tcaagttccg gggccacttc ctgatcgagg gcgacctgaa ccccgacaac
     1261 agcgacgtgg acaagctgtt catccagctg gtgcagacct acaaccagct gttcgaggaa
     1321 aaccccatca acgccagcgg cgtggacgcc aaggccatcc tgtctgccag actgagcaag
     1381 agcagacggc tggaaaatct gatcgcccag ctgcccggcg agaagaagaa tggcctgttc
     1441 ggcaacctga ttgccctgag cctgggcctg acccccaact tcaagagcaa cttcgacctg
     1501 gccgaggatg ccaaactgca gctgagcaag gacacctacg acgacgacct ggacaacctg
     1561 ctggcccaga tcggcgacca gtacgccgac ctgtttctgg ccgccaagaa cctgtccgac
     1621 gccatcctgc tgagcgacat cctgagagtg aacaccgaga tcaccaaggc ccccctgagc
     1681 gcctctatga tcaagagata cgacgagcac caccaggacc tgaccctgct gaaagctctc
     1741 gtgcggcagc agctgcctga gaagtacaaa gagattttct tcgaccagag caagaacggc
     1801 tacgccggct acatcgatgg cggagccagc caggaagagt tctacaagtt catcaagccc
     1861 atcctggaaa agatggacgg caccgaggaa ctgctcgtga agctgaacag agaggacctg
     1921 ctgcggaagc agcggacctt cgacaacggc agcatccccc accagatcca cctgggagag
     1981 ctgcacgcca ttctgcggcg gcaggaagat ttttacccat tcctgaagga caaccgggaa
     2041 aagatcgaga agatcctgac cttccgcatc ccctactacg tgggccctct ggccagggga
     2101 aacagcagat tcgcctggat gaccagaaag agcgaggaaa ccatcacccc ctggaacttc
     2161 gaggaagtgg tggacaaggg cgccagcgcc cagagcttca tcgagcggat gaccaacttc
     2221 gataagaacc tgcccaacga gaaggtgctg cccaagcaca gcctgctgta cgagtacttc
     2281 accgtgtaca acgagctgac caaagtgaaa tacgtgaccg agggaatgag aaagcccgcc
     2341 ttcctgagcg gcgagcagaa aaaagccatc gtggacctgc tgttcaagac caaccggaaa
     2401 gtgaccgtga agcagctgaa agaggactac ttcaagaaaa tcgagtgctt cgactccgtg
     2461 gaaatctccg gcgtggaaga tcggttcaac gcctccctgg gcacatacca cgatctgctg
     2521 aaaattatca aggacaagga cttcctggac aatgaggaaa acgaggacat tctggaagat
     2581 atcgtgctga ccctgacact gtttgaggac agagagatga tcgaggaacg gctgaaaacc
     2641 tatgcccacc tgttcgacga caaagtgatg aagcagctga agcggcggag atacaccggc
     2701 tggggcaggc tgagccggaa gctgatcaac ggcatccggg acaagcagtc cggcaagaca
     2761 atcctggatt tcctgaagtc cgacggcttc gccaacagaa acttcatgca gctgatccac
     2821 gacgacagcc tgacctttaa agaggacatc cagaaagccc aggtgtccgg ccagggcgat
     2881 agcctgcacg agcacattgc caatctggcc ggcagccccg ccattaagaa gggcatcctg
     2941 cagacagtga aggtggtgga cgagctcgtg aaagtgatgg gccggcacaa gcccgagaac
     3001 atcgtgatcg aaatggccag agagaaccag accacccaga agggacagaa gaacagccgc
     3061 gagagaatga agcggatcga agagggcatc aaagagctgg gcagccagat cctgaaagaa
     3121 caccccgtgg aaaacaccca gctgcagaac gagaagctgt acctgtacta cctgcagaat
     3181 gggcgggata tgtacgtgga ccaggaactg gacatcaacc ggctgtccga ctacgatgtg
     3241 gacgctatcg tgcctcagag ctttctgaag gacgactcca tcgataacaa agtgctgact
     3301 cggagcgaca agaaccgggg caagagcgac aacgtgccct ccgaagaggt cgtgaagaag
     3361 atgaagaact actggcgcca gctgctgaat gccaagctga ttacccagag gaagttcgac
     3421 aatctgacca aggccgagag aggcggcctg agcgaactgg ataaggccgg cttcatcaag
     3481 agacagctgg tggaaacccg gcagatcaca aagcacgtgg cacagatcct ggactcccgg
     3541 atgaacacta agtacgacga gaacgacaaa ctgatccggg aagtgaaagt gatcaccctg
     3601 aagtccaagc tggtgtccga tttccggaag gatttccagt tttacaaagt gcgcgagatc
     3661 aacaactacc accacgccca cgacgcctac ctgaacgccg tcgtgggaac cgccctgatc
     3721 aaaaagtacc ctaagctgga aagcgagttc gtgtacggcg actacaaggt gtacgacgtg
     3781 cggaagatga tcgccaagag cgagcaggaa atcggcaagg ctaccgccaa gtacttcttc
     3841 tacagcaaca tcatgaactt tttcaagacc gagattaccc tggccaacgg cgagatccgg
     3901 aagcggcctc tgatcgagac aaacggcgaa acaggcgaga tcgtgtggga taagggccgg
     3961 gactttgcca ccgtgcggaa agtgctgtct atgccccaag tgaatatcgt gaaaaagacc
     4021 gaggtgcaga caggcggctt cagcaaagag tctatcctgc ccaagaggaa cagcgacaag
     4081 ctgatcgcca gaaagaagga ctgggaccct aagaagtacg gcggcttcga cagccccacc
     4141 gtggcctatt ctgtgctggt ggtggccaaa gtggaaaagg gcaagtccaa gaaactgaag
     4201 agtgtgaaag agctgctggg gatcaccatc atggaaagaa gcagcttcga gaagaatccc
     4261 atcgactttc tggaagccaa gggctacaaa gaagtgaaaa aggacctgat catcaagctg
     4321 cctaagtact ccctgttcga gctggaaaac ggccggaaga gaatgctggc ctctgccggc
     4381 gaactgcaga agggaaacga actggccctg ccctccaaat atgtgaactt cctgtacctg
     4441 gccagccact atgagaagct gaagggctcc cccgaggata atgagcagaa acagctgttt
     4501 gtggaacagc acaaacacta cctggacgag atcatcgagc agatcagcga gttctccaag
     4561 agagtgatcc tggccgacgc taatctggac aaggtgctga gcgcctacaa caagcacaga
     4621 gacaagccta tcagagagca ggccgagaat atcatccacc tgtttaccct gaccaatctg
     4681 ggagcccctg ccgccttcaa gtactttgac accaccatcg accggaagag gtacaccagc
     4741 accaaagagg tgctggacgc caccctgatc caccagagca tcaccggcct gtacgagaca
     4801 cggatcgacc tgtctcagct gggaggcgac gcctatccct atgacgtgcc cgattatgcc
     4861 agcctgggca gcggctcccc caagaaaaaa cgcaaggtgg aagatcctaa gaaaaagcgg
     4921 aaagtggacg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
     4981 ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc
     5041 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
     5101 cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta ccccgaccac
     5161 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgcacc
     5221 atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
     5281 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
     5341 gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc cgacaagcag
     5401 aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag
     5461 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
     5521 aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac
     5581 atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga cgagctgtac
     5641 aagtaaccac cacactggac tagtggatcc gagctcggta ccaagcttaa gtttaaaccg
     5701 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     5761 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     5821 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     5881 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     5941 ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg
     6001 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     6061 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     6121 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     6181 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     6241 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     6301 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     6361 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg
     6421 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     6481 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     6541 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     6601 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     6661 atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag
     6721 tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc
     6781 attttcggat ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     6841 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     6901 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     6961 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg
     7021 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     7081 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     7141 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     7201 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     7261 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     7321 ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg
     7381 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     7441 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     7501 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     7561 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     7621 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     7681 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     7741 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta
     7801 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     7861 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     7921 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     7981 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     8041 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     8101 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     8161 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     8221 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     8281 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     8341 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     8401 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     8461 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     8521 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     8581 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     8641 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     8701 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     8761 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     8821 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     8881 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     8941 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     9001 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     9061 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     9121 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     9181 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     9241 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     9301 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     9361 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     9421 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     9481 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     9541 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     9601 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     9661 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     9721 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     9781 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     9841 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     9901 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     9961 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
    10021 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
    10081 ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga gatctcccga
    10141 tcccctatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagtatct
    10201 gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt aagctacaac
    10261 aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc gttttgcgct
    10321 gcttcgcgat gtacgggcca gatatacgcg tt
//
